Mitochondria are the primary sites of energy production in cells. Energy production occurs through the action of a series of enzyme complexes called the mitochondrial electron transport (or respiratory) chain. These complexes are responsible for: 1) the transport of electrons from NADH to oxygen and, 2) the coupling of oxidation to synthesis of ATP (oxidative phosphorylation). ATP then provides the primary source of energy for driving a cell's many energy-requiring reactions.
Most mitochondrial proteins are the products of nuclear genes and are imported into the mitochondria from the cytosol following their synthesis. Targeting of these proteins to mitochondria is achieved by an N-terminal leader (or signal) peptide of 10 to 70 amino acid residues which contains many positively charged amino acids. Once these precursor proteins are localized in the mitochondria, the leader peptide is cleaved by a signal peptidase to generate the mature protein. Most leader peptides are removed in a one step process by a protease termed mitochondrial processing peptidase (MPP) (Paces, V. et al. (1993) Proc. Natl. Acad. Sci. 90:5355-58). In some cases a two-step process occurs in which MPP generates an intermediate precursor form which is cleaved by a second enzyme, mitochondrial intermediate peptidase, to generate the mature protein.
MPP isolated from Neurospora crassa is a complex consisting of two dissociable components. The larger component has catalytic activity and has been called MPP; and the smaller component increases the activity of the complex and is called protease enhancing peptidase (PEP). MPP from Saccharomyces cerevisiae and rat liver is a heterodimer consisting of two dissimilar subunits, alpha (.about.55 kDa) and beta (.about.55 kDa) (Paces, et al. supra). In these species, it has not been determined whether the protease activity lies solely in the dimer or in a single subunit. The alpha-subunit of rat liver MPP is 36% identical to the large subunit (MPP) of N. crassa. The beta-subunit of rat liver MPP, beta-MPP, is most closely related to the smaller subunit of S. cerevisiae MPP (45% identity) and to N. crassa PEP (52% identity). Rat liver beta-MPP is also homologous (55% identity) to the core I protein from human ubiquinol-cytochrome c reductase (Hoffman, G. G. et al. (1993) J. Biol. Chem. 268:21113-19). Ubiquinol-cytochrome c reductase is one of the key enzyme complexes in the respiratory chain. It consists of 10-11 subunits, two of which are designated core I and core II proteins. The exact function of these two proteins is not known, but in yeast they appear to be necessary for assembly of the complex (Hoffman et al. supra). Although the core I protein of ubiquinol-cytochrome c reductase and PEP are identical proteins in N. crassa, the structurally homologous core I and beta-MPP proteins of yeast, rat, and human are genetically distinct.
Rat liver beta-MPP is 489 amino acids in length and is characterized by a 45-amino acid leader peptide that is positively charged and has a predicted signal peptidase cleavage site sequence, RST.sub.45 QA. Paces et al.(supra) suggest that, after being imported into the mitochondria, beta-MPP is cleaved by pre-existing MPP. An alpha-helical structure is predicted in the region between amino acids 165 and 205 of beta-MPP. The first part of this region, residues 168-186, is highly charged with several glutamate and arginine residues. This highly charged, amphiphilic helix, that is present in alpha-MPP as well, is believed to participate in binding of MPP to the positively charged, leader peptide substrates. An additional feature of beta-MPP and its homologs is the presence of one or two possible divalent metal-binding sites characteristic of metallopeptidases. Residues H.sub.101, E.sub.181, and H.sub.198 are believed to cooperate in binding of divalent metals such as Zn.sup.2+ or Mn.sup.2+, and the sequence H.sub.101 FLEH, unique to the rat liver beta-MPP, is also a potential metal-binding site (Paces, et al. supra).
The discovery of a new mitochondrial processing peptidase subunit and the polynucleotides encoding it satisfies a need in the art by providing new compositions which are useful in the diagnosis, prevention and treatment of smooth muscle disorders, neurological disorders, and cancer.